T cell tolerance to self-antigens and the regulation of immune responses to environmental antigens are an intrinsic part of the immune system. Immune tolerance is initially achieved during thymic development yet further regulation of autoreactive T cells is required in the periphery. This peripheral tolerance is achieved through a number of mechanisms, which include several types of CD4+ regulatory T cells. A defect in the function or generation of these regulatory T cells has been shown to lead to autoimmunity in mouse and man. We have shown that activation of human CD4+CD25- T cells leads to expression of both CD25 and FOXP3 and that these induced CD4+CD25+ T cells have regulatory properties. This suggests that in humans there is a mechanism by which CD4+CD25+FoxP3+ regulatory T cells can be generated in the periphery upon activation with the appropriate stimuli. In this grant we propose to pursue this finding and examine whether, in humans, induction of a CD4+CD25+FoxP3+ regulatory T cell (gTR) population in the periphery is a significant mechanism by which immune responses are regulated. We plan to address the hypothesis that in humans TR can be generated in the periphery using three specific aims: Aim 1. We hypothesize that during an inflammatory response a unique set of signals can drive the de novo generation of CD4+CD25+FoxP3+ TR from a pool of CD4+CD25- FoxP3- T cells. We propose, in this Aim, to identify the T cell compartment from which these regulatory T cells arise and examine the signals that lead to this differentiation and the requirements for survival and growth of the resulting gTR cells. Aim 2. We hypothesize that newly generated CD4+CD25+FoxP3+ regulatory T cells specific to self antigen arise during inflammation and that this is a mechanism by which autoreactive T cells activated at a site of inflammation and tissue injury are suppressed. We will approach this question in the context of T1D by evaluating whether islet specific antigens can lead to the de novo generation of TR and comparing the requirements for activation, survival, growth and resulting function of these islet specific TR to those derived from a foreign antigen. Aim 3. We hypothesize that a deficit in the ability to generate peripheral CD4 TR population is a key step which underlies human autoimmune diseases. In this Aim we will examine whether such a defect in the generation of TR is present in individuals with T1D and if that defect is specific to islet antigens.